DEVELOPING AN IN CELLULO PROTEIN CRYSTALLIZATION SYSTEM

Abstract

Developing an in cellulo protein crystallization system Kakanga Moses

Protein crystallization in living cells is often perceived as an atypical occurrence. Recently it was shown that p21 serine/threonine kinase 4 (PAK4) binds its inhibitor Inka1 to form rod like protein crystals whose X-ray structure was solved inside a single mammalian cell. The packing in these crystals consists of a hexagonal array with an 80 Å wide channel that allows inserting other proteins. This was tested by inserting GFP, which forms green crystals in mammalian cells. In this thesis we have further developed this system to accept larger proteins, increased the size of the crystals and accelerated the rate of crystal formation. We added inserts into the crystal scaffold to develop in vivo biosensors for visually monitoring protein–protein interactions. Thus, we conducted a proof of principle study that demonstrates that this system can be adapted for use in drug discovery and for screening inhibitors of protein–protein interactions.